scONE-seq: A one-tube single-cell multi-omics method enables simultaneous dissection of molecular phenotype and genotype heterogeneity from frozen tumors

Abstract Genomic and transcriptomic heterogeneity both play important roles in normal cellular function as well as in disease development. To be able to characterize these different forms of cellular heterogeneity in diverse sample types, we developed scONE-seq, which enables simultaneous transcriptome and genome profiling in a one-tube reaction. Previous single-cell-whole-genome-RNA-sequencing (scWGS-RNA-seq) methods require physical separation of DNA and RNA, often by physical separation of the nucleus from the cytoplasm. Most of these methods are labor-intensive and technically demanding, time-consuming, or require special devices, and they are not applicable to frozen samples that cannot generate intact single-cell suspensions. scONE-seq is a one-tube reaction which eliminates loss due to transfer steps, and thus is highly scalable and compatible with frozen biobanked tissue, generating data that is superior in quality compared to other applicable methods. We benchmarked scONE-seq against existing methods using cell lines and lymphocytes from a healthy donor, and we applied it to a 2-year-frozen astrocytoma sample profiling over 1,200 nuclei, subsequently identifying a unique transcriptionally normal-like tumor clone. scONE-seq makes it possible to perform large-scale single-cell multi-omics interrogation with ease on the vast quantities of biobanked tissue, which could transform the scale of future multi-omics single-cell cancer profiling studies.